Differentiation of division I (cfiA-negative) and division II (cfiA-positive) Bacteroides fragilis strains by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Abstract

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is increasingly used in clinical microbiological laboratories to identify bacteria and fungi at a species level and to subtype them. The cfiA gene encoding the unique carbapenemases found in Bacteroides is restricted to division II Bacteroides fragilis strains. The aim of this study was to evaluate whether MALDI-TOF MS is suitable for differentiating B. fragilis strains which harbour the cfiA gene from those that do not. A well-defined collection of 40 B. fragilis isolates with known imipenem MICs (0.062-.32 mg l "1 ) were selected for this study. Twelve B. fragilis strains with known cfiA status, including NCTC 9343 (division I) and TAL3636 (division II), were measured by means of microflex LT MALDI-TOF MS and well-defined differences in mass spectra between the cfiA-positive and cfiA-negative strains were found in the interval 4000-5500 Da. A further 28 strains were selected for the blind measurements: 9 cfiA-positive clinical isolates with different imipenem MICs ranging between 0.06 and .32 mg l "1 (different expressions of the metallo-b-lactamase gene) were clearly separated from the 19 cfiA-negative isolates. The presence or absence of the selected peaks in all tested strains clearly differentiated the strains belonging to B. fragilis division I (cfiA-negative) or division II (cfiA-positive). These results suggest a realistic method for differentiating division II B. fragilis strains (harbouring the cfiA gene) and to determine them at a species level at the same time. Although not all cfiA-positive B. fragilis strains are resistant to carbapenems, they all have the possibility of becoming resistant to this group of antibiotics by acquisition of an appropriate IS element for full expression of the cfiA gene, leading to possible treatment failure.

Biomedical Mass Spectrometry in Today's and Tomorrow's Clinical Microbiology Laboratories

“…Investigators noted a good correlation of MS types with cifA types based on the evolutionary history of the species. Apparently, the cifA gene has been acquired by one lineage and has not been further spread by horizontal gene transfer (25,33). Other resistance mechanisms that rely on the activity of single resistance determinants may be amenable to the same approach.…”

“…In the future, MS technology in clinical microbiology will be extended to include characterization of host and microbial proteins, sugars, fatty acids, nucleic acids, and metabolites in a single clinical sample (25). This will require greater integration into laboratory workflow and further technical improvement, especially with respect to analyte detection sensitivity.…”

“…Currently, the major commercial MALDI-TOF MS systems with validated microbial identification capacity are the Bruker Biotyper and the bioMérieux Vitek MS system (the latter developed by AnagnosTec [Germany] and Shimadzu Corporation [Japan]) (5,25). Andromas (Paris, France) provides an additional database that can be used in combination with the Bruker platform (2).…”

Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry-Based Functional Assay for Rapid Detection of Resistance against β-Lactam Antibiotics

“…Some automated systems can provide an antimicrobial susceptibility test (AST) report after a few hours. In the case of Bacteroides fragilis, it was recently shown that the differentiation between cfiA-negative and cfiA-positive isolates (cfiA encodes a class B metallo-beta-lactamase) was possible directly from matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) profile spectra (13,20). However, such a clonal differentiation of resistant and susceptible clones cannot be expected for the majority of bacteria.…”

A Novel Selective Medium for Isolation of Bacteroides fragilis from Clinical Specimens

“…Although MALDI-TOF MS has facilitated the identification of B. fragilis and its subsequent differentiation into division I and II strains, without a selective medium, recognition of B. fragilis in mixed growth with other bacteria remains difficult (9,13). Surveillance for drug-resistant B. fragilis strains in various patient populations also requires the availability of a selective and differential culture medium.…”